Transcriptomic responses in Japanese medaka (Oryzias latipes) exposed to 17β-estradiol

Abstract

The effects of 17β-estradiol (E2) were evaluated using the medaka DNA microarray representing 36,398 genes. We first evaluated chronic effects on medaka exposed to E2 at different concentrations for 60 days posthatch. At ≥ 30 ng/L of E2 severe reproductive impairments such as sex reversal were observed. Larval medaka, Oryzias latipes, (within 24 hrs posthatch) were then exposed to E2 at various concentrations (3, 30, 100 ng/L) for up to 7 days. Microarray analyses of the E2-exposed larvae revealed that exposure to E2 up-regulated and down-regulated 339 and 105 genes, respectively. The up-regulated genes included ones involved in the p53 signaling pathway, apoptosis, and growth and development, in addition to well-known biomarkers such as vitellogenin and choriogenins. Down-regulated genes included heat shock proteins and estrogen receptors. Most of the up-regulated genes encoding the p53 signaling pathway, apoptosis, and growth and development exhibited a dose-dependent increase in gene expression, whereas the down-regulated genes in the heat shock protein category showed a dose-dependent decrease in gene expression. Time course experiments suggested that the E2 treatment attenuated the time-dependent changes in gene expressions of these genes. Among the genes related to oocyte maturation, estrogen-regulated genes such as choriogenins and vitellogenins were dramatically induced in response to E2 exposure, whereas other steroid-regulated genes such as zona pellucida-domain proteins did not change in gene expression by the E2 treatment. Results suggest that transcriptomic studies on larval medaka help elucidate the effects caused by endocrine disruptors on various biological pathways in vertebrate development.

Species-specific considerations in using the fish embryo test as an alternative to identify endocrine disruption

“…Estrogens can pass the blood-brain barrier and have been shown to interfere with cholesterol synthesis and neurosteroidogenesis in vitro and in vivo in fish (Cypriani et al, 1988;Messa et al, 2005;Nagarajan et al, 2013;Sinchak et al, 2003;Smith et al, 1978), suggesting that embryonic steroidogenesis is likely to be influenced by some estrogenic chemicals. However, in this study it was found that 3ˇ-hsd (which encodes a downstream steroidogenic enzyme) was not affected by EE2 in medaka, supporting similar results for other steroidogenesis genes in medaka larvae and adults (Mizukami-Murata and Kishi-Kadota, 2011;Zhang et al, 2008). Therefore, the influence of estrogenic compounds on steroidogenesis appears complex.…”

“…Importantly, embryos of both species revealed endocrine responses specific to previously reported mode-of-action for the here-tested chemicals, and which were comparable to responses in larval and adult fish reported elsewhere (Leon et al, 2008;Mizukami-Murata and Kishi-Kadota, 2011;Zhang et al, 2008). While medaka provided a clearer and more chemical-specific picture of endocrine responses, zebrafish embryos were more sensitive than medaka embryos and showed a broader range of morphological effects.…”

“…The androgen receptor (ar) was not inhibited by EE2 in either 48-hpf or 96-hpf zebrafish embryos, whereas ar alpha was downregulated in medaka, as reported for medaka larvae and adult males elsewhere (Mizukami-Murata and Kishi-Kadota, 2011;Zhang et al, 2008). Because androgens are necessary for brain development, sexual differentiation and sexual behavior in male fish, the disruption of androgenic signaling in juveniles and adults can cause severe reproductive effects (Jobling et al, 2009;Leon et al, 2008;Oshima et al, 2003).…”

17β‐Trenbolone exposure programs metabolic dysfunction in larval medaka

“…The TB concentrations used were 0 (control), 2, 6, 20, 60, 100, and 200 ng/L (in dimethyl sulfoxide with a final concentration of 1:2500 v/v water). We previously demonstrated that highly reproducible gene expression data could be obtained from larval medaka through examining the variance in mRNA expression levels between pairs of control samples (0.95 average correlation coefficient among control samples; Mizukami‐Murata and Kishi‐Kadota, ). In addition, we demonstrated that at least 7 days of exposure are needed for the comprehensive analysis of gene responses to E2, because number of expression genes in larvae exposed for 1 and 2 days were significantly lower than that exposed for 7 days (Mizukami‐Murata and Kishi‐Kadota, ).…”

“…We previously demonstrated that highly reproducible gene expression data could be obtained from larval medaka through examining the variance in mRNA expression levels between pairs of control samples (0.95 average correlation coefficient among control samples; Mizukami‐Murata and Kishi‐Kadota, ). In addition, we demonstrated that at least 7 days of exposure are needed for the comprehensive analysis of gene responses to E2, because number of expression genes in larvae exposed for 1 and 2 days were significantly lower than that exposed for 7 days (Mizukami‐Murata and Kishi‐Kadota, ). Based on the results of the preceding physiological experiments, we conducted a microarray analysis of larval medaka that were exposed to 0–200 ng/L TB for 7 d. Each TB treatment contained 90 larvae in each chamber.…”